This project aims to investigate the process of vacuole formation (endocytosis) in human erythrocytes and in their plasma membranes (erythrocyte ghosts). The main goals are to investigate the mechanism of the process of endocytosis, and to compare endocytosis in erythrocyte ghosts from diseased patients with normal erythrocyte ghosts. In particular, ghosts from patients with hereditary spherocytosis and Kx deficiency disease will be studied. Endocytosis depends on a supply of ATP both in whole erythrocytes and in ghosts. In ghosts, and probably in whole erythrocytes as well, the ATP must be hydrolyzed for endocytosis to occur. This probably occurs through the agency of one of the Mg ion-ATPases or Ca ion-Mg ion ATPases. The project proposes to investigate the nature of these ATPases, and the relationship between them and endocytosis in ghosts. Pure erythrocyte ghosts appear to behave differently from whole erythrocytes with regard to the effects of Ca ion on endocytosis. If the motile system of ghosts is analogous to the contractile system of muscle, this difference is probably due to regulatory proteins which have been removed from the pure ghosts. A preparation of crude ghosts or whole erythrocytes will be made which consistently shows these differences, and then location or isolation of the regulatory factors will be attempted. The effects on endocytosis of cations such as Mg ion and Ca ion will be investigated, and compared with the effects of such cations on non-energy dependent shape changes such as the discocyte to echinocyte transition. The effects of inactivating agents will be observed, to see whether the Mg ion-ATPase or the Ca ion-Mg ion ATPAse is inactivated along with endocytosis.